1、Designation:D781712(Reapproved 2021)Standard Test Method forEnumeration of Yeast and Mold in Raceway Brine,Brine-Cured Hides and Skins1This standard is issued under the fixed designation D7817;the number immediately following the designation indicates the year oforiginal adoption or,in the case of r
2、evision,the year of last revision.A number in parentheses indicates the year of last reapproval.Asuperscript epsilon()indicates an editorial change since the last revision or reapproval.1.Scope1.1 This test method covers the enumeration of yeast andmold.This test method is applicable to raceway brin
3、e,brine-cured hides and skins,and pre-charge raceway liquor.1.2 The values stated in SI units are to be regarded asstandard.No other units of measurement are included in thisstandard.1.3 This standard does not purport to address all of thesafety concerns,if any,associated with its use.It is therespo
4、nsibility of the user of this standard to establish appro-priate safety,health,and environmental practices and deter-mine the applicability of regulatory limitations prior to use.1.4 This international standard was developed in accor-dance with internationally recognized principles on standard-izati
5、on established in the Decision on Principles for theDevelopment of International Standards,Guides and Recom-mendations issued by the World Trade Organization TechnicalBarriers to Trade(TBT)Committee.2.Referenced Documents2.1 ASTM Standards:2D6715 Practice for Sampling and Preparation of Fresh orSalt
6、-Preserved(Cured)Hides and Skins for Chemical andPhysical TestsE691 Practice for Conducting an Interlaboratory Study toDetermine the Precision of a Test MethodE177 Practice for Use of the Terms Precision and Bias inASTM Test Methods3.Summary of Test Method3.1 Samples of brine-cured hides and skins,r
7、aceway brine,or pre-charge raceway liquor are serially diluted and plated onagar containing 7%NaCl and an antibiotic solution.The platesare incubated at 20 25 C for 5 days.4.Significance and Use4.1 This test method enumerates salt tolerant yeast andmold,and under the conditions of this test method t
8、hose areequated as halophilic organisms.Salt tolerant yeast and moldhave been known to cause damage to hides and skins inraceway brine.5.Apparatus5.1 Incubator,20 25 C.5.2 Colony counter(not mandatory,but highly recom-mended).5.3 Sterile pipets.5.4 Stomacher,for mixing initial dilution.(If stomacher
9、 isunavailable,hand-mix.)5.5 Balance.5.6 Sterile petri dishes.5.7 Autoclave(sterilizer)(Check the effectiveness of ster-ilization weekly.For example,place spore suspensions or stripsof Bacillus stearothermophilus(commercially available)insideglassware for a full autoclave cycle.Follow manufacturersd
10、irections for sterilization of specific media.)5.8 pH meter.5.9 Waterbath,45 6 1 C.5.10 Stomacher bags,or sterile,sealable quart plastic bag(for example,food storage type,sterile bag).5.11 Cutting tool,sterile(for example,scalpel blade andforcep,as needed for cutting cured hides and skins).5.12 Vort
11、ex mixer,for mixing dilution tubes(optional).5.13 Autoclave thermometer.6.Reagents and Materials6.1 Butterfields Phosphate Stock SolutionDissolve 34 gKH2PO4(Potassium Phosphate monobasic)in 500 mL DIwater.Adjust the pH to 7.2 6 0.1 with 1N 6N NaOH.Bringvolume to 1 L with DI water.Sterilize for 15 mi
12、n at 121 C.1This test method is under the jurisdiction of ASTM Committee D31 on Leatherand is the direct responsibility of Subcommittee D31.02 on Wet Blue.Current edition approved Sept.1,2021.Published October 2021.Originallyapproved in 2012.Last previous edition approved in 2016 as D7817 12(2016).D
13、OI:10.1520/D7817-12R21.2For referenced ASTM standards,visit the ASTM website,www.astm.org,orcontact ASTM Customer Service at serviceastm.org.For Annual Book of ASTMStandards volume information,refer to the standards Document Summary page onthe ASTM website.Copyright ASTM International,100 Barr Harbo
14、r Drive,PO Box C700,West Conshohocken,PA 19428-2959.United StatesThis international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for theDevelopment of International Standards,Guides and Recommendations is
15、sued by the World Trade Organization Technical Barriers to Trade(TBT)Committee.1NOTE1Typical autoclave setting is 120 124 C.(See 5.7.)6.2 Butterfields Phosphate Diluent with salt(BPD w/salt)Take 1.25 mL of Butterfields Phosphate Stock solution(6.1)and bring to 1 L with DI water,then add 77 g of salt
16、(NaCl)perlitre prior to autoclaving.Dispense into 1-L bottles and 9-mLdilution tubes.Sterilize for 15 min at 121 C.(See Note 1.)6.3 Potato Dextrose Agar(PDA).6.4 Antibiotic solution(Chloramphenicol)3(needed toinhibit bacterial growth on agar).6.5 Distilled or deionized water.6.6 Salt(NaCl),Sodium chloride reagent grade.6.7 1N 6N NaOH.6.8 Bacillus stearothermophilus spore suspensions or strips(commercially available),or equivalent.7.Hazards7.1 All reagents and chemicals should be handled with car
